JP7264540B2 - Composition for prevention or treatment of asthma, rhinitis or conjunctivitis containing an N-acyl-amino acid as an active ingredient - Google Patents

Description

The present invention relates to a preventive or therapeutic composition for asthma, rhinitis or conjunctivitis, containing an N-acyl-amino acid as an active ingredient.

Asthma is an inflammatory disease of the respiratory tract that is caused by genetic and environmental factors and is a condition in which the bronchi in the lungs become hypersensitive, i.e., the narrowing of the bronchi often makes it difficult to breathe, accompanied by an audible wheeze. It is a disease that presents with severe coughing symptoms. In recent years, there is a tendency to increase the occurrence of environmental pollution and exposure to chemicals. Asthma occurs in all age groups, but about 30% of children are children. 17(5):881-6).

Rhinitis is a disease that exhibits an inflammatory reaction in the nasal mucosa, and can be broadly classified into allergic rhinitis and non-allergic rhinitis. Allergic rhinitis appears variously depending on the specific type, but mainly shows symptoms such as sneezing, runny nose, stuffy nose, and mucosal cell dilatation due to specific immunoglobulin (IgE, IgM, etc.) abnormal reaction. Non-allergic rhinitis includes, in addition to coryza due to viral infections, infectious rhinitis due to bacterial and fungal infections, vasomotor rhinitis that can occur from abnormalities of the autonomic nervous system distributed in the nasal mucosa, use of incompatible drugs, cold There are physical causes such as temperature, rhinitis caused by food, and rhinitis caused by structural abnormality of the nasal cavity.

Worldwide, the prevalence of allergic rhinitis continues to increase. Already in the 20th century, direct medical costs (outpatient costs, prescriptions and emergency room visits) for allergic rhinitis in the United States exceeded $1.9 billion, with additional losses due to complications amounting to $4 billion. According to the health insurance statistics chronology in 2011, allergic rhinitis surged from 2 million in 2000 to 5.6 million in 2011 (from 14th to 5th) in the ranking of outpatient diseases among Koreans. As a result, it ranked 4th among the 500 types of diseases in terms of salaries paid by the National Health Insurance Corporation. It is thought that the social cost of allergic rhinitis will continue to increase in the future, and the trend is to increase the demand for treatment or amelioration methods for allergic rhinitis.

Conjunctivitis is a disease in which an allergic reaction occurs in the conjunctiva, and is frequently accompanied by allergic diseases such as hay fever and allergic rhinitis. It is a typical seasonal disease, and symptoms worsen in spring, but improve in autumn and winter. It is mainly seen in people with allergic predispositions, and it begins before puberty and recurs for 5 to 10 years. However, recently, it is known that people of all ages can get sick regardless of the season or physical constitution while abnormally high temperatures continue. Pollutants such as exhaust gas and chemical dust are also known to have a large impact. Common symptoms are very itchy eyes, severe redness and eyelid edema, along with profuse tearing, which can be made worse by rubbing the eyes. There may also be scratching or burning pain, foreign body sensation, blindness, and subconjunctival hemorrhage.

Asthma treatment mainly relies on drug therapy, and in addition to anti-inflammatory treatment (oral administration and inhalation of steroids), bronchodilators (theophylline) and leukotriens inhibitors as a type of anti-inflammatory agent are generally used. (Clin Exp Allergy. 2012 42:650-8).

For allergic rhinitis, the use of antihistamines is recommended to alleviate symptoms, but high doses of antihistamines may induce drowsiness and dizziness, and long-term use may lead to excessive secretion of histamine, resulting in symptoms. It can get even more intense.

Steroidal anti-inflammatory drugs of the glucocorticoid family, which are representative therapeutic agents for conjunctivitis, are commonly used as anti-inflammatory drugs and show excellent effects on rheumatoid arthritis, etc. It suppresses or prevents various inflammatory reactions, including immune stimulation.

Steroids are currently the most widely used drugs for allergic inflammatory diseases, including asthma, rhinitis and conjunctivitis. In the case of asthma, acute asthma is known to be relatively well regulated by steroids. However, many patients with chronically advanced asthma or severe asthma do not respond to steroids, and many patients with such symptoms have been suffering for a long time without any treatment, and are unable to live their lives. quality is markedly reduced, leading to cruel death (Clin Exp Allergy. 2012 42:650-8).

A problem with steroid use is that steroids cause serious side effects. Steroids have fatal side effects that decrease immune response and increase microbial infection and cancer development, and function of skin, muscle, bone, eye, nervous system, endocrine system, cardiovascular system, immune system, fire extinguishing system, etc. It leads to abnormalities and other diseases, and its use is very limited (N Engl J Med. 2005, 353:1711-23).

Therefore, the development of a safe anti-allergic anti-inflammatory drug that can replace steroids is one of the most important and urgent problems to be solved by modern medicine. Researchers in this field have developed non-steroidal anti-inflammatory drugs (NSAIDs) to overcome the side effects of steroids. Aspirin is a typical example, and aspirin is an enzyme involved in prostaglandin synthesis from arachidonic acid produced by the action of cytosolic phospholipase A2 (cPLA ₂ ). COX) inhibitors, and besides aspirin, many NSAIDs (Indomethacin, Ibuprofen, Celecoxib, Rofecoxib, etc.) are COX inhibitors. However, these NSAIDs are not used for the treatment of allergic inflammatory diseases due to their weak anti-inflammatory action compared to steroids. Moreover, scientists have been developing p38 mitogen-activated protein kinase and _cPLA2 inhibitors, which are inflammatory response-inducing enzymes, for quite some time, but these drugs are currently in clinical use due to side effects. Currently, there is no (Bioorg Med Chem 2008; 16: 1345-58).

In this way, the development of ``steroid adjuvant drugs'' and ``steroid alternative drugs'' is thought to make a significant contribution to the treatment of various allergic inflammatory diseases, including asthma. there is

Therefore, the present inventors have made intensive research efforts to develop a drug that has no side effects and is excellent in improving asthma, rhinitis and conjunctivitis. , confirmed that it has an inhibitory effect on rhinitis and conjunctivitis, and completed the present invention.

Accordingly, an object of the present invention is to provide a composition for preventing, improving or treating asthma containing an N-acyl-amino acid as an active ingredient.

Another object of the present invention is to provide a composition for preventing, improving or treating allergic rhinitis containing an N-acyl-amino acid as an active ingredient.

Still another object of the present invention is to provide a composition for preventing, improving or treating conjunctivitis containing an N-acyl-amino acid as an active ingredient.

Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

According to one aspect of the present invention, the present invention provides a composition for preventing, improving or treating asthma, rhinitis or conjunctivitis containing an N-acyl-amino acid as an active ingredient.

According to another aspect of the invention, the invention provides a method of preventing, ameliorating or treating asthma, rhinitis or conjunctivitis comprising administering to a subject an effective amount of an N-acyl-amino acid.

According to still another aspect of the present invention, the present invention provides uses of N-acyl-amino acids for the prevention, amelioration or treatment of asthma, rhinitis or conjunctivitis.

In the present invention, asthma is a disease in which the bronchial tubes in the lungs are extremely sensitive, that is, the bronchial tubes are often narrowed, making it difficult to breathe and causing severe coughing accompanied by wheezing, It is an allergic disease caused by allergic inflammation of the bronchi.

In the present invention, rhinitis is a disease that causes an inflammatory reaction in the nasal mucosa, and can be broadly classified into allergic rhinitis and non-allergic rhinitis. Allergic rhinitis is a hypersensitivity reaction of the nasal mucosa to specific substances. After the substances (antigens) that cause allergy are exposed to the nasal mucosa, mast cells, eosinophils, and other cells are formed in the stimulated area. It means a disease in which inflammatory cells mediated by various IgE antibodies gather and inflammatory reactions occur due to various mediators secreted by them. Non-allergic rhinitis includes, in addition to coryza due to viral infections, infectious rhinitis due to bacterial and fungal infections, vasomotor rhinitis that can occur from abnormalities of the autonomic nervous system distributed in the nasal mucosa, use of incompatible drugs, cold There are physical causes such as temperature, rhinitis caused by food, and rhinitis caused by structural abnormality of the nasal cavity.

In the present invention, the conjunctivitis means conjunctivitis caused by local sensation, ranging from only mild hyperemia of the conjunctiva to eyelid swelling and conjunctival edema. A type of allergy is called allergic conjunctivitis.

The N-acyl-amino acids of the present invention are preferably N-acyl-alanine or N-acyl-tryptophan, more preferably N-acyl-L-alanine or N-acyl-L-tryptophan.

The N-acyl-L-alanine or N-acyl-L-tryptophan is a form in which the alpha amino group of alanine or tryptophan is acylated, and may be synthesized by an organic synthesis method, using commercially available reagents. You can buy it and use it.

As used herein, the term "acyl" or "acyl group" is not particularly limited and refers to the remaining atomic groups of a carboxylic acid excluding OH, which is the carboxy group, and is generally represented by RCO. R is one or more substituents, and is not limited as long as it is a substituent capable of bonding to CO. When the above R is an aromatic atomic group, it may be particularly referred to as "aroyl", which is also a kind of acyl group. Examples of said acyl include formyl (HCO-), acetyl (CH ₃ CO-), propionyl (C ₂ H ₅ CO-), butyryl (C ₃ H ₇ CO-), valeryl (C ₄ H ₉ CO-). , pentanoyl (CH ₃ (CH ₂ ) ₃ CO—), palmitoyl (C ₁₅ H ₃₁ CO—), stearoyl (C ₁₇ H ₃₃ CO—), oleoyl (C ₁₇ H ₃₁ CO—), oxalyl (—CO— CO—), malonyl (—COCH ₂ CO—), succinyl (—CO(CH ₂ ) ₂ CO—), benzoyl (C ₆ H ₅ CO—), toluoyl (CH ₃ —C ₆ H ₄ —CO—), salicylyl (HO--C ₆ H ₄ --CO--), cinnamoyl (C ₆ H ₅ CH=CHCO--), naphthoyl (C ₁₀ H ₇ CO--), phthaloyl (CO--C ₆ H ₄ --CO--), furoyl ( C ₅ H ₃ O ₂ —), undecanoyl (CH ₃ (CH ₂ ) ₉ CO—), docosenoyl obtained by removing the OH group from docosenoic acid, but not limited thereto.

Said N-acyl-alanine is preferably, but not limited to, N-acetyl-alanine or N-oleoyl-alanine .

Said N-acyl-tryptophan is preferably, but not limited to, N-acetyl-tryptophan or N-oleoyl-tryptophan .

The N-acyl-L-alanine is preferably, but not limited to, N-acetyl-L-alanine or N-oleoyl-L- Alanine (N-oleoyl-L-alanine).

The N-acyl-L-tryptophan is preferably, but not limited to, N-acetyl-L-tryptophan or N-oleoyl-L- Tryptophan (N-oleoyl-L-tryptophan).

On the other hand, in the present specification, the terms "containing as an active ingredient" or "effective amount" mean containing an amount sufficient to achieve the efficacy or activity of the N-acyl-amino acid. In one embodiment of the present invention, the N-acyl-amino acid in the composition of the present invention is, for example, 10 μg/kg or more, preferably 0.1 mg/kg or more, more preferably 1 mg/kg or more, still more preferably 10 mg/kg. /kg or more. Since N-acyl-amino acids have almost no adverse effects on the human body even when administered in an excessive amount, the upper limit of the amount of N-acyl-amino acids contained in the composition of the present invention can be selected within an appropriate range by those skilled in the art. can be implemented.

The N-acyl-amino acid used as an active ingredient in the composition of the present invention is not only the compound itself, but also its pharmaceutically, food or cosmetically acceptable salts, hydrates and solvates. are interpreted to include products or prodrugs.

As used herein, the terms "pharmaceutically acceptable salt", "foodstuff acceptable salt" or "cosmetically acceptable salt" do not severely irritate the organism to which the compound is administered. means a dosage form of a compound that does not provoke or impair the biological activity and physical properties of the compound. The pharmaceutically-, food- or cosmetically-acceptable salts can be obtained by adding the compounds of the present invention to inorganic acids such as hydrochloric acid, bromic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p - sulfonic acids such as toluenesulfonic acid, tartaric acid, formic acid, citric acid, acetic acid, trichloroacetic acid, trifluoroacetic acid, capric acid, isobutanoic acid, malonic acid, succinic acid, phthalic acid, gluconic acid, benzoic acid, lactic acid, fumaric acid , maleic acid, salicylic acid, and the like. Alternatively, the compound of the present invention may be reacted with a base to form a salt such as an ammonium salt, an alkali metal salt such as sodium or potassium salt, an alkaline earth metal salt such as calcium or magnesium salt, dicyclohexylamine, N-methyl-D-glucamine, It can also be obtained by forming salts with organic bases such as tris(hydroxymethyl)methylamine, and amino acids such as arginine and lysine, without limitation.

As used herein, the terms "pharmaceutically acceptable hydrate", "food acceptable hydrate" or "cosmetically acceptable hydrate" refer to Refers to hydrates of said N-acyl-amino acids that have physical effects. The terms "pharmaceutically acceptable solvate", "foodstuff acceptable solvate" or "cosmetically acceptable solvate" refer to compounds that have the desired pharmacological effect. Refers to the solvates of said N-acyl-amino acid compounds. Said hydrates and solvates can also be prepared using said acids and are broadly included in said pharmaceutically, food or cosmetically acceptable salts.

As used herein, the terms “pharmaceutically acceptable prodrug”, “food acceptable prodrug” or “cosmetically acceptable prodrug” refer to the N-acyl-amino acid Refers to derivatives of said N-acyl-amino acids that must undergo biotransformation before exerting their pharmacological effects. Such prodrugs are manufactured for prolonged duration of action and reduced side effects to improve chemical stability, patient acceptability, bioavailability, organ selectivity or convenience of preparation. . The production of the prodrug of the present invention is carried out using the N-acyl-amino acid according to a conventional method in the art (eg, Burger's Medicinal Chemistry and Drug Chemistry, 5th ed., 1: 172-178 and 949-982 ( 1995)).

According to a preferred embodiment of the invention, the composition of the invention is a pharmaceutical composition.

According to a preferred embodiment of the invention, the pharmaceutical composition of the invention comprises a pharmaceutically acceptable carrier.

Pharmaceutically acceptable carriers contained in the pharmaceutical composition of the present invention are those commonly used in formulations, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate and gelatin. , calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, talcum, magnesium stearate and mineral oil. isn't it. The pharmaceutical composition of the present invention may further contain lubricating agents, wetting agents, sweetening agents, flavoring agents, emulsifying agents, suspending agents, preservatives, etc., in addition to the above ingredients. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

The pharmaceutical composition of the present invention can be administered orally or parenterally. In the case of parenteral administration, nasal administration, eye drop administration, intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, percutaneous administration, rectal administration. It can be administered by injection, intradural administration, intracerebrovascular injection, and the like.

Appropriate doses of the pharmaceutical composition of the present invention include formulation method, administration mode, patient age, body weight, sex, disease state, diet, administration time, administration route, excretion rate and reaction sensitivity. A physician of ordinary skill can readily determine and prescribe an effective dosage for the desired treatment or prevention. According to a preferred embodiment of the invention, the daily dosage of the pharmaceutical composition of the invention is 10 μg-1,000 mg/kg.

The pharmaceutical composition of the present invention is formulated using a pharmaceutically acceptable carrier and/or excipient by a method that can be easily carried out by a person having ordinary knowledge in the technical field pertaining to the present invention. It may be manufactured in unit dose form by packaging, or may be manufactured in multi-dose containers. At this time, the dosage form may be in the form of a solution, suspension or emulsion in an oil or aqueous medium, or may be in the form of an extract, powder, granules, tablets or capsules, and may be dispersed or stabilized. It may further contain an agent.

The pharmaceutical composition of the present invention can be prepared in the form of skin external preparations, aerosols, sprays, eye drops, oral preparations and injections.

The pharmaceutical composition of the present invention can be used alone for the prevention and treatment of asthma, rhinitis or conjunctivitis, or in combination with methods such as surgery, radiotherapy, hormone therapy, chemotherapy and biological response modifiers. can do.

The pharmaceutical composition of the invention may be used for humans or animals.

As used herein, the term "subject" or "subject" means a human or animal in need of administration of said N-acyl-amino acid.

As used herein, the term "prevention" means any action of suppressing or delaying progression of asthma, rhinitis or conjunctivitis by administration of the composition of the present invention.

As used herein, the term "treatment" means any action in which asthma, rhinitis or conjunctivitis is ameliorated or beneficially altered by administration of the compositions of the present invention.

According to a preferred embodiment of the invention, the composition of the invention is a food composition.

The food composition according to the present invention can be used in functional foods or added to various foods. Foods to which the composition of the present invention can be added include, for example, beverages, alcoholic beverages, confectionery, diet bars, dairy products, meats, chocolate, pizza, breads, ramen, other noodles, gums, and ice creams. , vitamin complexes, and health supplements.

The food composition of the present invention can contain not only the N-acylamino acid as an active ingredient, but also ingredients normally added during food production, such as proteins, carbohydrates, fats, nutrients, seasonings and flavoring agents. . Examples of carbohydrates mentioned above are monosaccharides, such as glucose, fructose, etc.; disaccharides, such as maltose, sucrose, oligosaccharides, etc.; Sugar alcohols such as erythritol. Natural flavors [thaumatin, stevia extracts (eg rebaudioside A, glycyrrhizin, etc.]) and synthetic flavors (saccharin, aspartam, etc.) can be used as flavoring agents. For example, when the food composition of the present invention is produced as drinks and beverages, in addition to N-acyl amino acids, citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, and various plants Extracts and the like can further be included.

In addition to the above, the food composition of the present invention contains various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH. Modifiers, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated beverages, and the like may be included. In addition, the composition of the present invention can contain pulp for the production of natural fruit juices, fruit juice beverages and vegetable beverages. Such ingredients may be used individually or in combination. The proportion of such additives is not very critical, but is generally chosen in the range of 0.01 to 10 parts by weight per 100 parts by weight of the composition of the invention.

The present invention provides a food with health claims as a food composition containing an N-acyl-amino acid or a food-acceptable salt thereof as an active ingredient. Functional health foods refer to foods that have bioregulatory functions such as disease prevention and improvement, bioprotection, immunity, recovery after illness, and anti-aging, and should be harmless to the human body when taken for a long time. The health functional food is a food produced by adding N-acyl-amino acid to food materials such as beverages, teas, spices, gums, confectionery, or by encapsulating, powdering, suspension, etc. It is a substance that shows specific effects on health when ingested, but unlike general medicines, it has the advantage that it does not have side effects that may occur when taking medicine for a long time because it is made from food. The food with health claims of the present invention thus obtained is very useful because it can be ingested on a daily basis. The amount of N-acyl-amino acid to be added to such food with health claims varies depending on the type of the food with health claims, and cannot be specified uniformly, but may be added within a range that does not impair the original taste of the food. , usually in the range of 0.01 to 50% by weight, preferably 0.1 to 20% by weight, relative to the target food. In addition, in pill, granule, tablet or capsule type health functional foods, it is usually added in the range of 0.1 to 100% by weight, preferably in the range of 0.5 to 80% by weight. In one specific example, the functional health food of the present invention can be in the form of pills, tablets, capsules or beverages.

As used herein, the term "amelioration" means any action in which general symptoms associated with asthma, rhinitis, or conjunctivitis do not progress or improve as a result of ingestion or administration of the composition of the present invention.

The food composition of the present invention may be used as a human food or animal feed or feed additive.

According to a preferred embodiment of the invention, the composition of the invention is a cosmetic composition.

When the composition of the present invention is produced as a cosmetic composition, the composition of the present invention contains not only the above-described N-acyl-amino acids, but also ingredients commonly used in cosmetic compositions, such as antioxidants. agents, stabilizers, solubilizers, vitamins, the usual adjuvants such as pigments and perfumes, and carriers. In addition, the composition of the present invention may be prepared by mixing a conventionally used ameliorating agent or sedative for asthma, rhinitis or conjunctivitis, in addition to the N-acyl-amino acid described above, to the extent that the action is not impaired. can be used.

As the carrier, purified water, monohydric alcohols (ethanol or propyl alcohol), polyhydric alcohols (glycerol, 1,3-butylene glycol or propylene glycol), higher fatty acids (palmitic acid or linolenic acid), oils ( Wheat germ oil, camellia oil, jojoba oil, olive oil, squalene, sunflower oil, macadamia nut oil, avocado oil, hydrogenated soybean lecithin or fatty acid glycerides) can be used, but not limited thereto. In addition, surfactants, bactericides, antioxidants, ultraviolet absorbers, anti-inflammatory agents and cooling agents can be added as necessary.

Surfactants include polyoxyethylene, hydrogenated castor oil, polyoxyethylene, oleyl ether, polyoxyethylene monooleate, polyoxyethylene, glyceryl monostearate, sorbitan monostearate, polyoxyethylene monooleate, sorbitan, It can be selectively included from the group consisting of sucrose fatty acid ester, hexaglyceryl monolaurate, polyoxyethylene-reduced lanolin, POE, glycerylpyroglutamic acid, isostearic acid, diester, N-acetylglutamine and isostearyl ester.

Bactericides can be selectively included from the group consisting of hinokitiol, triclosan, chlorhexidine gluconate, phenoxyethanol, resorcinol, isopropylmethylphenol, azulene, salicylic acid and zinc pyrithione.

Any of butylhydroxyanisole, acetic acid, propyl acetic acid and erythorbic acid may be used as the antioxidant.

UV absorbers include benzophenones such as dihydroxybenzophenone, melanin, ethyl para-aminobenzoate, para-dimethylaminobenzoic acid 2-ethylhexyl ester, cinoxate, para-methoxycinnamic acid 2-ethylhexyl ester, 2-(2-hydroxy-5-methylphenyl ) Any of benzotriazole, urocanic acid and metal oxide fine particles may be used.

Dipotassium glycyrrhizinate or allantoin can be used as an antiphlogistic agent, and red pepper tincture or 1-menthol can be used as a cooling agent.

The dosage form of the composition is any dosage form that can incorporate an N-acyl-amino acid as an active ingredient, such as tonic water, shampoo, rinse, hair conditioner, hair spray, powder, gel, cream, essence, lotion, sol-gel. , emulsion, oil, wax, spray, mist, etc., but not limited to these.

A summary of the features and advantages of the present invention are as follows:
(i) The present invention provides compositions for preventing, improving or treating asthma, rhinitis or conjunctivitis using N-acyl-amino acids.

(ii) The composition of the present invention can be usefully used to safely and effectively improve or treat allergic diseases and asthma caused by various causes without worrying about side effects.

Photographs showing inhibition of inflammation on H&E staining of lung tissue upon oral administration of N-acyl-L-alanine, N-oleyl-L-alanine, N-acyl-L-tryptophan, and N-oleyl-L-tryptophan ( OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan, NOT = N-oleyl-L-tryptophan). On masson's trichrome staining of lung tissue upon oral administration of N-acyl-L-alanine, N-oleyl-L-alanine, N-acyl-L-tryptophan, N-oleyl-L-tryptophan Photographs showing inhibition of bronchial fibrosis (OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan, NOT = N-oleyl -L-tryptophan). Fig. 2 shows the inhibitory effect of airway cell-derived Th2 cytokine (IL4) secretion of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan ( * p<0.05 vs yolk albumin group; OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan, NOT = N- oleyl-L-tryptophan). Fig. 2 shows the inhibitory effect of airway cell-derived Th2 cytokine (IL5) secretion of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan ( * p<0.05 vs yolk albumin group; OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan, NOT = N- oleyl-L-tryptophan). Fig. 2 shows the inhibitory effects of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan on airway cell-derived Th2 cytokine (IL13) secretion ( * p<0.05 vs yolk albumin group; OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan, NOT = N- oleyl-L-tryptophan). Fig. 6A is a diagram showing the rhinitis reaction inhibitory effects of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan (Fig. 6A: nose rubbing action, Fig. 6B: sneezing) (*p<0.05 vs yolk albumin group; OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan , NOT=N-oleyl-L-tryptophan). Fig. 6A is a diagram showing the rhinitis reaction inhibitory effects of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan (Fig. 6A: nose rubbing action, Fig. 6B: sneezing) (*p<0.05 vs yolk albumin group; OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan , NOT=N-oleyl-L-tryptophan). FIG. 7 shows the inhibitory effect of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan on allergic conjunctivitis (FIG. 7A: eyeball photograph, diagram 7B: Clinical grade) (*p<0.05 vs yolk albumin group; OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N- Acetyl-L-tryptophan, NOT=N-oleyl-L-tryptophan). FIG. 7 shows the inhibitory effect of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan on allergic conjunctivitis (FIG. 7A: eyeball photograph, diagram 7B: Clinical grade) (*p<0.05 vs yolk albumin group; OVA = yolk albumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N- Acetyl-L-tryptophan, NOT=N-oleyl-L-tryptophan).

The present invention will be described in more detail below using examples. Those of ordinary skill in the art should understand that these examples are merely for the purpose of more specifically illustrating the invention and that the scope of the invention is not limited by the spirit of the invention. will be clear to those

[Example]
[Preparation example 1. Preparation of experimental animals]
BALB/c mice (6-week-old, male) free of specific pathogens were purchased from Doal Biotech (Osan, Korea). Mice were used at the start of the experiment, aged 7-10 weeks, housed in a laminar flow cabinet and provided with water and food ad libitum.

[Preparation example 2. Preparation of drugs to be used]
The drugs N-acetyl-L-alanine, N-acetyl-L-tryptophan, etc. used in this patent were purchased from Sigma (Missouri, USA) and N-oleyl-L-alanine was purchased from Cayman (Michigan, USA). state). N-oleyl-tryptophan was synthesized by the following method. First, to produce methyl oleyl-L-tryptophanate, oleic acid (2.36 mmol), L-tryptophan methyl ester hydrochloride (2.60 mmol), 1-ethyl-3-(3-dimethylaminopropyl) A mixture of carbodiimide (EDCI) (2.60 mmol), hydroxybenzotriazole (HOBt) (2.60 mmol) and triethylamine (11.8 mmol) dissolved in dichloromethane was stirred at room temperature for 12 hours. The reaction was concentrated, diluted with saturated NaHCO ₃ solution, and then extracted with ethyl acetate three times. The extracted organic solvent layer was collected, washed with brine, washed with 1N HCl, and then washed with brine again. The organic solvent layer was dried over anhydrous MgSO ₄ , concentrated and then purified on medium pressure liquid chromatography (MPLC) using n-hexane and ethyl acetate. The yield was 73% and the properties of methyloleyl-L-tryptophanate are as follows.

¹ H NMR (500 MHz, Chloroform-d) δ 8.18 (s, 1H), 7.56 (dd, J = 7.9, 1.0 Hz, 1H), 7.39 (dt, J = 8.1, 0.9 Hz, 1H), 7.29 (s , 1H), 7.22 (ddd, J = 8.2, 7.1, 1.2 Hz, 1H), 7.14 (ddd, J = 8.0, 7.0, 1.0 Hz, 1H), 7.00 (d, J = 2.4 Hz, 1H), 5.99 ( d, J = 7.9 Hz, 1H), 5.37 (qd, J = 4.1, 2.1 Hz, 2H), 5.00 (dt, J = 8.0, 5.3 Hz, 1H), 3.72 (s, 3H), 3.39 - 3.28 (m , 2H), 2.20 - 2.13 (m, 2H), 2.08 - 1.99 (m, 5H), 1.60 (t, J = 7.4 Hz, 2H), 1.34 - 1.28 (m, 24H), 0.92 - 0.89 (m, 3H ). LC-MS ₍ ESI) _, calcd for _{C30H46N2O3 482.4} , found m/z 483.4 (M+H ⁺ ).
To produce N-oleyltryptophan from methyloleyl-L-tryptophanate, NaOH (1.48mmol) was added to a solution of methyloleyl-L-tryptophanate (0.37mmol) in tetrahydrofuran, and the mixture was stirred at room temperature. and stirred for 12 hours. Water was added to the reactant and extracted with dichloromethane. After adjusting the pH to 1 by adding 1N HCl to the aqueous layer, it was extracted three times with ethyl acetate. The extracted organic solvent layer was dried over anhydrous _MgSO4 and concentrated. The yield was 86%, and the properties of N-oleyltryptophan are as follows.
¹ H NMR (500 MHz, Chloroform-d) δ 8.22 (s, 1H), 7.61 (d, J = 7.9 Hz, 1H), 7.40 (dt, J = 8.1, 0.9 Hz, 1H), 7.23 (ddd, J = 8.1, 6.9, 1.2 Hz, 1H), 7.15 (ddd, J = 8.0, 7.1, 1.0 Hz, 1H), 7.08 (d, J = 2.4 Hz, 1H), 5.37 (qd, J = 5.3, 4.6, 2.3 Hz, 2H), 4.95 (dt, J = 7.3, 5.6 Hz, 1H), 3.45 - 3.33 (m, 2H), 2.17 - 2.10 (m, 2H), 1.59 - 1.51 (m, 2H), 1.38 - 1.26 ( m, 26H) _, 0.91 (d, J = 6.8 Hz, 3H). _{C29H44N2O2 468.3} , found _m /z 469.3 (M + H ⁺ ).

[Preparation example 3. mouse asthma induction]
Mice were sensitized by intraperitoneal injection twice on days 0 and 14 with 0.02 mg ovalbumin (OVA) and alum (1 mg) as an adjuvant (Sigma-Aldrich). For 4 days from the 28th day, the mice were placed in plastic nebulizer cylinders of a certain size, and 5% OVA was sprayed with oxygen gas for 30 minutes, and the mice were allowed to inhale OVA in the form of aerosol. This was named 'airway challenge'.

[Preparation example 4. mouse rhinitis induction]
Mice were sensitized by two intraperitoneal injections of 0.02 mg ovalbumin (OVA) and alum (1 mg) as an adjuvant (Sigma-Aldrich) intraperitoneally on days 0 and 14. . From the 21st day for 7 days, 100 μg of OVA was dissolved in 20 μl of phosphate buffered saline once a day, and 10 μl of each was injected into both nasal cavities for local stimulation. After the last intranasal OVA instillation (experimental day 27), the sum of the number of nasal rubbing and sneezing caused by mice is defined as the symptom score. Symptom scores were recorded for 15 minutes to compare the results of each experimental group, and the results are shown in FIG.

[Preparation example 5. Induction of conjunctivitis in mice]
Mice were sensitized by two intraperitoneal injections of 0.5 mg ovalbumin (OVA) and alum (1.5 mg) (Sigma-Aldrich) as an adjuvant intraperitoneally on days 0 and 7. made. For 12 days from the 14th day, 100 μg of OVA was dissolved in 10 μl of phosphate buffered saline once a day, and 10 μl of the solution was injected into both eyes for local stimulation to induce conjunctivitis.

[Preparation example 6. Measurement of inflammatory cells in bronchi]
Mice are anesthetized, a tube is inserted into the bronchi, and the bronchi are washed with buffer to obtain cells. This was centrifuged and suspended at 1×10 ⁴ cells/100 μl, centrifuged and fixed on a slide using a cytospin, and Diff-Quik staining revealed eosinophils, monocytes, neutrophils and large cells. Measure the number of phagocytic cells.

[Preparation example 7. Lung tissue staining]
Lungs are excised for histological analysis, fixed in formalin, and embedded in paraffin. The embedded tissue is sectioned to a thickness of 4 μm and 1) stained with H/E (Hematoxylin and Eosin) to examine the degree of inflammation, and 2) stained with Masson's trichrome to examine the degree of lung fibrosis.

[Preparation example 8. Clinical grading measurement of conjunctival edema and redness]
Clinical grading of conjunctival edema and redness was performed 24 hours after the final OVA instillation (experimental day 25) by portable slit lamp, Cail Zeiss, Oberkochen, Germany. rice field. Evaluations were performed in a blinded fashion 2 hours after OVA injection. Each clinical parameter was determined according to Merayo-Lloves, J. Med. , Calonge, M, and Foster, C.; S. 1995. Experimental model of allergic connectivity to ragweed in guinea pig. Curr. Eye Res. 14:487-494, ranging from 0 to 4+ (0 absent, 1+ minimal, 2+ slight, 3+ moderate, 4+ severe). One group received no treatment as a negative control group.

[Example 1. Asthma inhibitory effect of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan]
[Example 1-1. Suppressive effect of administration of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan on airway inflammatory cell influx in asthma-induced mice]
Neutrophils, macrophages, monocytes and eosinophils were observed as airway inflammatory cells. N-Acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, N-oleyl-L-tryptophan, each corresponding to 3.5 μmole per kg, was added to 10 ml of phosphoric acid. It was dissolved in buffered saline and administered orally once a day for 7 days starting on the 25th day. As can be seen in Table 1, oral administration of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, N-oleyl-L-tryptophan caused intra-airway Both significantly inhibited the influx.

OVA = ovalbumin, NAA = N-acetyl-L-alanine, NOA = N-oleyl-L-alanine, NAT = N-acetyl-L-tryptophan, NOT = N-oleyl-L-tryptophan, MAC = macrophages, NEU=neutrophils, MONO=monocytes, EOS=eosinophils, N=number of mice, S.E. D. = standard deviation, Min = minimum value, Max = maximum value [Example 1-2. Degree of inflammation on H&E staining of lung tissue]
H&E staining of lung tissue was performed after one week of oral drug administration. In the normal group (normal), there were almost no inflammatory cells around the airways, and in the OVA-challenged group (OVA), many inflammatory cells were extensively gathered around the airways, and the bronchi were narrow. , whereas peribronchial angiogenesis can be observed (Fig. 1), oral administration of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, N-oleyl-L-tryptophan In the groups (OVA+NAA, OVA+NOA, OVA+NAT, OVA+NOT), the degree of inflammation was remarkably suppressed, and the effect of N-oleyl-L-alanine was particularly suppressed (Fig. 1).

[Example 1-3. Measurement of Cytokines in Bronchial Lavage]
Multiple cytokine (TNF-α, IL-4, IL-5, IL-13) concentrations in bronchial lavage fluid were measured using ELISA kits (R&D Systems, Minneapolis, Minn, USA).

[Example 2. Bronchial closure inhibitory effect of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan]
Masson's trichrome staining of lung tissue was performed after one week of oral drug administration. In the normal group (normal), there was almost no fibrosis around the airways, and in the OVA-challenged group (OVA), a large degree of fibrosis was observed around the airways (Fig. 2). -L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan orally administered groups (OVA+NAA, OVA+NOA, OVA+NAT, OVA+NOT) show such a degree of fibrosis. Although it was suppressed, the action of N-oleyl-L-alanine in particular was the most suppressed (Fig. 2).

[Example 3. N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, N-oleyl-L-tryptophan airway cell-derived Th2 cytokine (IL4, IL5 and IL13) secretion inhibitory effect]
The Th2 cytokines IL-4, IL-5 and IL-13 are known to be involved in airway inflammation, bronchial hyperresponsiveness and IgE antibody production through eosinophils. N-Acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, N-oleyl-L-tryptophan, each corresponding to 3.5 μmole per kg, was added to 10 ml of phosphoric acid. It was dissolved in buffered physiological saline and orally administered once a day for 7 days from the 25th day after induction of asthma, and cytokines were measured. Oral administration of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan and N-oleyl-L-tryptophan significantly increased IL-4, IL-5 and IL-13. (Figs. 3, 4 and 5).

[Example 4. Sinusitis reaction inhibitory effect of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan]
Ovalbumin (OVA) of Preparative Example 3 was used as an antigen for inducing allergic rhinitis, and the drug was administered from day 21 to day 26 of the test 12 hours after OVA challenge. -L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan and N-oleyl-L-tryptophan were each dissolved in phosphate buffered saline to 10 µM, and 10 µl of each was administered into the nasal cavity. After the last intranasal OVA instillation (experimental day 27), the total number of nasal rubbing and sneezing by mice was defined as a symptom score and measured. Symptom scores were recorded for 15 minutes to compare the results of each experimental group, and the results are shown in FIG. In the normal group, there was little nose rubbing or sneezing, and in the OVA-challenged group there was a marked increase in nose rubbing or sneezing (FIGS. 6A and 6B), whereas N-acetyl-L-alanine, N Nasal rubbing and sneezing frequency were significantly reduced in all groups to which oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan were intranasally administered (FIGS. 6A and 6B).

[Example 5. Conjunctivitis inhibitory effect of N-acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan]
The yolk albumin of Preparation 4 was used as the antigen for inducing conjunctivitis. N-Acetyl-L-alanine, N-oleyl-L-alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan were each dissolved in phosphate-buffered saline at 100 µM, and 10 µl was added to each eye. was administered to On day 26, clinical grading of conjunctival edema and redness was performed using a portable slit lamp, Cail Zeiss, Oberkochen, Germany, for 24 hours following ovalbumin injection. went later. Clinical grade compared the results of each experimental group, and the results are shown in FIG. The normal group had few symptoms of conjunctivitis, and the group challenged with yolk albumin alone had an increased clinical grade of conjunctivitis (FIGS. 7A and 7B), whereas N-acetyl-L-alanine, N-oleyl-L- The clinical grade was significantly decreased in all of the groups that received alanine, N-acetyl-L-tryptophan, and N-oleyl-L-tryptophan eye drops (FIGS. 7A and 7B).

Although specific parts of the present invention have been described in detail above, for those skilled in the art, such specific descriptions are merely preferred embodiments, and the scope of the present invention is not limited thereto. The point is clear. Accordingly, the substantial scope of the invention will be defined by the appended claims and their equivalents.

Claims (2)

A pharmaceutical composition for preventing or treating asthma, rhinitis or conjunctivitis, comprising an N-acyl-amino acid or a pharmaceutically acceptable salt thereof as an active ingredient ,
The N-acyl-amino acid is N-acetyl-alanine, N-oleoyl-alanine, N-acetyl-tryptophan, or N - a pharmaceutical composition which is N-oleoyl-tryptophan . A food composition for preventing or improving asthma, rhinitis or conjunctivitis, comprising an N-acyl-amino acid or a pharmaceutically acceptable salt thereof as an active ingredient ,
The N-acyl-amino acid is N-acetyl-alanine, N-oleoyl-alanine, N-acetyl-tryptophan, or N - a food composition which is N-oleoyl-tryptophan .

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